Background: Breast carcinigenesis results from changes in gene expression, primary activation of oncogenes and down-regulation of tumour suppressor genes. One of recently discovered genes involved in breast carcinogenesis is WW domain-containing oxidoreductase (WWOX) located on chromosome 16q23.3-24.1. Aim: The aim of the present study was to elucidate the role of tumour suppressor gene WWOX in normal and pathological cell metabolism. Materials and methods: The research was carried out on normal immortalized human mammary epithelial cell line HBL-100 and malignant human mammary epithelial cell line MDA-MB-231. Both cell lines were transduced with WWOX cDNA (WWOXw-t, WWOXΔ6-8 and WWOXantisense) cloned into pLNCX2 retroviral vector. Ectopic WWOX expression was confirmed by means of real-time RT-PCR. The transduced cells were further subjected to the following analyses: soft agar growth assay, Matrigel invasion assay, cell growth in Matrigel, and cell cycle analysis by cytometry. Results and conclusions: The results of our research demonstrated that an increase in WWOX expression in both cell lines dramatically decreased their anchorage-dependent growth in soft agar. For the colonies with diameter from 100 to 400 µm the average number of colonies formed by MDA-MB-231/vector was 127±4, for MDA-MB-231/WWOXΔ6-8 it was 65±21, whereas for MDA-MB-231/WWOXw-t it was only 9±2. In the case of normal cells (HBL-100) the average number of colonies with diameter 100-400 µm was 1669±43 for HBL-100/vector, while for HBL-100/WWOXw-t it was only 660±77. On the contrary to the results obtained in the soft agar assay, Matrigel invasion assay showed that an increase in WWOX expression resulted in higher cell migration through basement membrane. Both MDA-MB-231 and HBl-100 cell lines transduced with WWOXw-t cDNA exhibited branching morphogenesis, unlike the cells transduced with empty vector only, which formed spheroids. On the basis of those results, one might presume that WWOX is probably involved in formation of mammary gland structures. Flow cytometric analysis of DNA content revealed that no significant changes in cell cycle were observed in both cell lines transduced either with vector or with WWOX cDNA. Increased aggressiveness had no influence on cell cycle.

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